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Laboratory of Human Nutrition and Clinical Research Center Massachusetts Institute of Technology, Cambridge, MA 02139; and Nutrition Unit, Department of Medical Sciences, Uppsala University, Uppsala, SE-752–37 Sweden
1To whom correspondence should be addressed.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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KEY WORDS: • urea • nitrogen • leucine oxidation • balance • young adults
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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, the results of which have been confirmed in many
subsequent investigations (Walser 1981
), it has been
recognized that the rate of urea production (UP) generally exceeds its
rate of excretion via the kidneys. This is attributed to the hydrolysis
of urea by the resident microflora within the intestinal lumen, which
can be virtually eliminated by appropriate antibiotic treatment
(Walser and Bodenlos 1959
). The fates of the urea N,
released as ammonium N, are multiple, including fecal N loss
(Moran and Jackson 1990
), incorporation into bacterial
protein (Metges et al. 1999
), loss as ammonia in
feces (Wrong et al. 1985
), absorption across the
intestinal wall and incorporation into the metabolic N (amino acid)
pool (Giordano et al. 1968
, Metges et al. 1996
, 1999
, Niiyama et al. 1979
) and/or its direct return to the urea cycle
(Jackson et al. 1984
, Long et al. 1978). Clearly, therefore, urea is not merely an inert
product of tissue and organ N metabolism but may be utilized and so
contribute further to the economy of whole body N metabolism.
Jackson (1992)
considers that the N entering body tissues via urea
hydrolysis makes an important contribution to the achievement of N
balance, even at adequate N intakes. Importantly, he has concluded that
there is a control over urea hydrolysis which accounts for maintenance
of a constant UP rate, over about a four-fold range of intakes
(60–250 mg N · kg-1 · d-1)
(Jackson 1998
,1999
). We have questioned
the conclusion that UP is relatively constant, since we have observed
in one recent study (Forslund et al. 1998
) a parallel
increase in urea production when healthy adults were given two levels
of dietary protein within the supramaintenance range of total intake.
Hence, it was important to evaluate Jackson’s proposal in some further
detail and, in particular, his premise that there is no simple
relationship between UP and N intake over a wide range of protein
intakes which are sufficient to sustain N balance (Jackson 1998
). Also, Jackson (1998)
has suggested that the rate of urea
excretion is not a measure of amino acid oxidation. A previous study by
El-Khoury et al. (1994)
does not support this suggestion, and the
results of a recent study on urea kinetics from Jackson’s group
(Child et al. 1997
) also reveal that in men, but not in
women, N intake and UP were correlated. However, this latter study did
not control for intake since it was conducted in subjects, of different
body weight and composition, who were consuming their free-choice
intakes.
We previously conducted a series of studies of urea kinetics in subjects who were first adjusted to diets supplying different, but constant, levels of N intake. Hence, we thought it worthwhile to bring together the results of these different investigations, all using an identical, 24-h tracer approach with the purpose of determining the rates of urea production and of leucine oxidation, as an index of whole-body amino acid oxidation. Our analysis of these studies is the focus of this report.
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METHODS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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Data for this study have been drawn from four investigations, conducted in our laboratories over the past 5 y. All of the studies received approval from the relevant ethics committees of the respective institutions where the studies were carried out. All subjects gave their written informed consent and were paid for their participation. The details of two of these studies have been published, as provided below. Therefore, the designs of the two as yet unpublished experiments (expts.1 and 2) are presented along with a synopsis of the two published experiments (expts. 3 and 4), as follows:
Expt. 1: (Protein-free diet study).
Young adults (n = 11) participated in this
investigation, with 6 subjects (5 males, 1 female) studied in one group
(age: 23 ± 2 y; weight 63 ± 10 kg; height 169 ± 7 cm) and five males in another group (age 27 ± 6 y; weight
71 ± 14 kg; height 178 ± 5 cm). This was an investigation
of obligatory oxidative amino acid losses (OOAL), involving two 5-d
experimental diet periods and with one being a protein-free diet.
The other period was also 5-d and involved either giving a
leucine-free or sulfur amino acid-free, but otherwise adequate
diet. Data for this second period are not relevant to the present
investigation and will be published separately, with reference to the
assessment of OOAL. One of these groups, received, during the
protein-free diet period, therefore, an infusion of
[1-13C]leucine, and for these subjects the
leucine oxidation data are included here in our analysis of N
intake-urea kinetic relationships. The protein-free diet
supplied a daily energy intake that was constant for each subject,
ranging from 41 to 45 kcal · kg–1
(172–188kJ · kg–1) and was provided as
40% from fat (safflower oil and butter) and 60% from carbohydrate
(beet sugar and wheat starch). Vitamins, minerals, choline and fiber
(20 g of microcrystalline cellulose) were supplied as daily supplements
to meet or exceed recommended intakes (FNB, 1989
), and
as described elsewhere (El-Khoury et al. 1994
).
On the afternoon of d 5 of the protein-free diet period, subjects were admitted to the infusion room of the MIT Medical Department. After collecting blood and breath samples to measure background 13C isotopic enrichments, at just before 1800 h, priming boli of [13C]sodium bicarbonate (0.8 µmol · kg-1) (MassTrace, Woburn, MA), [15N,15N]urea (88 µmol · kg-1) (CIL, Cambridge, MA) and (for 5 of the 11 subjects) L-[1-13C]leucine (4.2 µmol · kg-1) (CIL) were administered. Then, the [15N,15N]urea (7 µmol · kg-1 · h-1) and [1-13C]leucine (2.8 µmol · kg-1 · h-1) tracers were infused continuously throughout a 24-h period, terminating at 1800 h on d 6.
Three meals, each providing one-third of the daily intake, were given at 2000, 0600 and 1200 h (noon).
Expt. 2 (Low-protein diet study).
Four male subjects participated in this experiment (age 22 ± 3 y; weight 73 ± 18 kg; height 175 ± 7 cm). Each was
given for 14 d a low-N diet based on an L-amino
acid mixture that was patterned after the indispensable amino acid
profile of hen’s egg protein (El-Khoury et al. 1998
),
except for its leucine content which was adjusted to approximate that
of mixed proteins in the body (Widdowson et al. 1979
).
The diet was otherwise similar to that for expt. 1 and provided an
energy intake of ~45kcal ·
kg-1 · d-1(188
kJ · kg-1 · d-1)
to maintain body weight.
On the afternoon of d 13 a 24-h tracer study began at about
1800 h; details are given elsewhere (El-Khoury et al. 1994
). L-[1-13C]leucine and
[15N,15N]urea were given
as intravenous tracers, as described above, and the tracer protocol was
terminated at 1800 h on the following day.
Expt. 3 ("Normal"- protein intake).
This study, involving seven adult male subjects, has been described in
detail elsewhere (El-Khoury et al. 1994
). The
experimental diet, based on egg protein (El-Khoury et al. 1998
), supplied 1.0 g
protein · kg-1 · d-1
and ~188 kJ · kg-1 ·
d-1. Leucine intake was 80
mg · kg-1 · d-1
and an additional 9.4
mg · kg-1 · d-1
was given as tracer during the 24-h tracer study. During the first
6 d, subjects were given three meals per day; the last meal on d 6
was given at 1500 h and the 24-h
[15N,15N]urea and
[1-13C]leucine tracer study started at
1800 h, lasting until 1800 h the next day (d 7). The feeding
regimen on d 7 consisted of hourly, small meals, to achieve a steady
metabolic fed state. The remaining details of the diet and tracer
infusion protocol were all as described previously (El-Khoury et al. 1994
) and similar to those for expts. 1 and 2 above.
Expt. 4 ("Normal-" and "High-" protein intake). Healthy male volunteers (n = 8) [age 27 ± 14 (means ± SD) y; weight 78 ± 7 kg, height 187 ± 6 cm] participated in the normal protein intake study and 6 healthy male volunteers [age 27 ± 15 (means ± SD) y; weight 80 ± 12 kg, height 186 ± 9 cm) in the high-protein intake study. One person participated in both studies, but urea kinetic results for another subject given the normal protein level were not included in this summary because of a suspected error in the amount of 15N-urea tracer administered. The subjects were recruited from the population of students and employees at Uppsala University.
A standardized diet was consumed during the 7-d experimental period, which was based on two major components: i) a milk drink as the principal protein source, flavored with banana or raspberry and ii) specially prepared cookies as an energy source to balance energy expenditure. During the experiment, at the "normal" protein intake, the diet supplied 1g · kg-1 · d-1; during the "high" protein intake it was 2.5 g · kg-1. Milk protein, from skim milk powder, was the principal protein source in both and the fat/carbohydrate energy ratio was kept at 40:60 for both diets. The dietary carbohydrate sources were wheat starch, sucrose and lactose. Cellulose powder (25 g) was included in each diet. Energy intake was given to maintain body energy balance.
The subjects were studied on an outpatient basis during d 1–5 at the
Energy Metabolic Unit (UPPCAL) of the Department of Nutrition, Uppsala
University. They were given the experimental diets for 7 d, and a
standard physical exercise program during each day was performed using
a cycle ergometer unit. During d 1–5, the food was given as three
major meals (breakfast, lunch and dinner) with two small meals in
between. During d 6 and 7, the food was equally distributed as 10 small
hourly meals from 1200 h until 2100 h. On d 7, as previously
described (Forslund et al. 1998
), a tracer/metabolic
study was conducted, involving a continuous infusion of
13C-leucine and 15N,15N-urea. A
detailed description of the procedures and of the experimental design
has been presented (El-Khoury et al. 1997
,
Forslund et al. 1998
).
Samples and analytical procedures
Details of the standard 24-h tracer-infusion protocol, used
in all of the above experiments, have been previously described
(El-Khoury et al. 1994
, 1997
), including
intravenous infusion techniques, blood and breath sampling, indirect
calorimetry and collection/analysis of plasma samples for
13C-
-ketoisocaproic acid
[13C-KIC] and
[15N,15N]urea, as well as
methods to account for breath
13CO2 "background"
enrichment and [13C]bicarbonate recovery.
Plasma and urinary urea N concentrations were determined by means of a
modified version of the procedure of Marsh et al. (1965)
. Urinary urea
excretion was corrected for the changes in body urea pool, as
previously described (El-Khoury et al. 1994
). Total
urinary N concentration was determined by micro-Kjeldahl analysis
(El-Khoury et al. 1994
).
Evaluation of data
The results have been summarized as means ± SD The principal statistical procedure used for evaluation of the data sets was linear regression. Complete data (urea N production, hydrolysis and excretion and leucine oxidation) were not available for all 36 subjects for various reasons, and so the number of data sets (varying from 29 to 34) used in the regression analyses are indicated in the figures below. Only 29 subjects were studied for leucine oxidation and for two subjects UP rates were not obtained. In one case this was because the urea pool was not primed with 15N,15N-urea and in the other case the plasma urea data were highly variable, possibly due to a malfunctioning of the infusion pump. Hence, UP rates are reported for a total of 34 subjects.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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and 2
the mean data obtained in each experiment, for urea kinetics and
metabolism and for leucine oxidation and balance, respectively, are
summarized. In Figure 1
the relationship between the level of N intake and rate of urea N
production is shown, indicating a highly positive linear relationship
(r = 0.98; P < 0.01) between these
variables. Similarly, urea N hydrolysis increased linearly with N
intake (r = 0.7; P < 0.05), but there
was considerable variation in the rates of hydrolysis among individual
subjects, especially at the normal protein intake. From these data
(Tables 1
and 2
and Fig. 1
) a number of points can be made: urea N
production and urea hydrolysis each appear to be a linear function of
the level of N intake. At low N intakes the amount of urea hydrolyzed
was low, amounting to 8 and 22 mg urea
N · kg-1 · d-1
for the protein-free and low-protein diets, respectively (Table 1)
. At the high N intake (392 mg
N · kg-1 · d-1),
the absolute rate of urea N hydrolysis was about four times that for
the low-protein diet. This compares with the nine-fold
difference in the level of the N intake between these two diets. It
should also be noted that while there was a significant linear
relationship between N intake and urea hydrolysis, it is possible there
is an upper rate of hydrolysis and that this was approximated within
the range of adequate and high protein intake studied. Only additional
studies, including studies at higher protein intakes, will answer this
question.
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The daily rate of leucine oxidation changed in parallel with
the level of leucine intake (Fig. 2
; Table 2
). Furthermore, leucine oxidation was highly correlated with
the rates of urea N production (Fig. 3
) and urea N excretion (Fig. 4
). At the low-protein and normal-protein intakes in expts. 2 and
3, respectively, the mean ratio of the rate of leucine oxidation to
urea N production (wt/wt) was 0.53 and 0.57, whereas at the
high-protein intake it was 0.65. Similarly, the mean ratios of the
rate of leucine oxidation to urea N excretion were 0.69, 0.73 and 0.83,
respectively. The difference between these ratios for the low and
higher protein intakes is probably due, in large part, to the higher
dietary leucine-to-nitrogen intake ratio in those given the
high-protein intakes (Table 2)
. Finally, it might also be noted
that the slope of the relationship between leucine oxidation and N
excretion (i.e., 1.17; Fig. 4
) was less than that between oxidation and
N production (i.e., 1.5; Fig. 3
). This, presumably, is due to the fact
that UP exceeds urea excretion and to a relatively greater degree at
normal and high vs. low-protein intakes.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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), in which
healthy adults were given a diet supplying protein at supramaintenance
levels equivalent to 1 and 2.5 protein
kg-1day-1, we observed that
the rates of UP and excretion changed in parallel with the level of
protein intake. This finding is contrary to the conclusion drawn by
Jackson (1998)
, that the UP rate is relatively constant over a wide
range of protein (N) intakes. However, Jackson’s studies have not
formally included as many observations as we have made at the higher
intake levels of protein. Jackson (1998)
has also proposed that a
control of body N homeostasis is achieved via altered rates of urea
hydrolysis, such that the rate of UP is relatively constant over a wide
range of intakes from inadequate to adequate levels and that urea
hydrolysis is increased to promote body protein (N) retention under
conditions of inadequate protein intakes. Because the conclusions and
interpretations drawn by Jackson were based on data for intakes that
did not include as high a range of intake as that studied recently by
us (Forslund et al. 1998
), we considered it worthwhile
to examine, in the aggregate, results obtained from a number of
comparable studies carried out in our laboratories on urea and leucine
kinetics. The measurements were made in subjects receiving different
intakes that ranged from an essentially protein (N) free intake through
maintenance to a high-intake level well beyond that needed for
maintenance of body protein balance.
From the present analysis, clearly the rates of urea N production and
excretion reflect the rate of amino acid (leucine) oxidation, and our
study questions the generality of Jackson’s (1998)
view that:
"... it is unsafe to presume that the rate of urea excretion is a
measure of the rate of urea production and hence of the rate of amino
acid oxidation." Our analysis supports the conclusion that we drew
earlier concerning the value of a measure of whole body leucine
oxidation to predict irreversible protein N loss (IPNL) or whole body
amino acid oxidation (El-Khoury et al. 1994
). It should
be noted that IPNL can only be determined precisely from leucine
oxidation rates when the dietary leucine-to-N intake ratio approximates
that of whole-body mixed proteins.
It is not entirely clear why our database and analysis lead to such a
different picture of urea N metabolism and kinetics in comparison with
that obtained from the extensive series of careful studies conducted by
Jackson and his colleagues, both in England and Jamaica. The tracer
model that these investigators have most frequently used involves a
primed/intermittent intravenous or oral administration of
15N15N urea over periods of
24 to 48 h (Jackson et al. 1984
), with analysis of
the 15N in urinary urea by isotope ratio mass
spectrometry and assessment by a model which includes use of the
appearance of both double and single
15N-labeled urea species in the specimens.
His group has also used a protocol (Child et al. 1997
,
Jackson et al. 1993
), involving a single dose of
[15N,15N]urea given
orally, with the amount of label excreted as
[15N,15N]urea and
[15N,14N]urea in urine
over the subsequent 48 h measured. In our view, both protocols
permit a reliable estimate of the steady-state rate of urea N
production (UP) and of urea N hydrolysis (H), with the latter
calculated as the difference between UP and urea N excretion. Jackson,
in his various studies, also uses the information on the excretion of
the double
([15N,15N]urea) and
single ([15N,14N]urea)
labeled species to determine the fate of the hydrolyzed urea N, either
with respect to its use in amino acid and protein synthesis or urea
resynthesis. Although we (El-Khoury et al. 1996
,
Forslund et al. 1998
) have raised questions about the
extent to which N exchange might complicate the estimates of the net
fate of hydrolyzed urea N, this issue is not critical here since we are
only concerned with the assessment of results for UP and hydrolysis.
Thus, we conclude that a reliable estimate of the rate of UP can be
achieved given an accurate determination of the total excretion of urea
N and of the double labeled species of 15N urea.
The urea supplementation studies by Meakins and Jackson (1996)
confirm
the adequacy of their primed/intermittent oral dosing protocol for
assessment of the rate of urea appearance (production). Similarly, the
24-h tracer protocol and plasma isotope dilution technique that we have
used in the present study for measurement of the rate of UP has been
validated (El-Khoury et al. 1994
), although we recognize
that it may underestimate acute changes in the rate of UP in response
to altered substrate availability. This is because of the large body
urea pool with a relatively slow turnover rate (Hamadeh and Hoffer 1998
). Again this is also not an important technical
issue for the present purposes. In summary, therefore, it does not seem
to us that the differences noted above between our experiments and
conclusions and those of and by Jackson and his colleagues
(Jackson 1998
) are due importantly to experimental
differences in the specific 15N- tracer protocols
applied in our respective investigations. A closer analysis of the
findings reported in a number of the earlier publications is,
therefore, desirable.
Thus, Child et al. (1997)
studied urea kinetics, based on a single oral
dose of 15N,15N-urea,
in healthy and light men whose free-choice N intake between the two
groups varied considerably, and they found that the rate of UP
correlated with N intake. This was not the case for light and heavy
women, probably because there was a much narrower difference in the
habitual level of protein intakes between the two groups of females.
Similarly, the results by Danielson and Jackson (1992)
and Meakins and Jackson (1996)
show that the rates of UP vary with the level of N
intake. In our judgment, therefore, these findings do not seem to offer
strong support for the proposition that i) UP tends toward a
constant relationship with body size or fat-free mass and
ii) within an adequate range of protein intakes UP does not
relate to intake of protein (Jackson 1998
,
1999
). However, Langran et al. (1992)
did not find a
difference in the UP rate when subjects received 75 or 35g protein/d
each for 5 d and so this does support Jackson’s proposition
(1998). Nevertheless, these latter findings should be evaluated with
due regard to the fact that subjects consuming the 75 g daily
protein level were in marked positive N balance. This would not be
expected for the intake level and may suggest an undercollection of
excreta and of N loss and which could explain the findings. However,
for whatever the reason, the results of this study by Langran et al. (1992)
appear to be at variance to those cited above from the same
laboratory.
Additionally, a number of Jackson’s studies have shown, again in our
judgment, that the rate of urea hydrolysis correlates positively with N
intake (Danielsen and Jackson 1992
, Meakins and Jackson 1996
) although this has not been a consistent finding
among their investigations (Jackson et al. 1990
,
Langran et al. 1992
). We see an apparently wide
variation in urea hydrolysis, especially at the more usual or
"normal" level of protein intake. Hence, comparisons made of this
parameter of urea kinetics among different studies, both within and
among laboratories, should be made with caution.
In similar studies, the question of the possible influence of length of
the adaptation or adjustment period, prior to conduct of the kinetic
studies, might be raised. We think that this is not an important
problem with respect to the comparisons drawn here because we have
previously shown (Rand et al. 1976
, Young 1999
) that by about 4–5 d a new and relatively
steady-state of N excretion and N balance is achieved. Also, in the
present low-protein study (El-Khoury, A. E. & Young, V. R., unpublished data, 1999) we found that urea excretion reached an
essentially constant rate after 4 d and that was sustained through
d 12.
In conclusion, we have found a distinct and essentially linear
relationship between protein intake, UP and hydrolysis and amino acid
(leucine) oxidation. Thus, our findings do not support the novel
concept elaborated by Jackson (1998
, 1999)
that a key control of N
balance and retention is exerted via a regulation of urea hydrolysis,
with subsequent incorporation of the ammonia released into -amino N
in the endogenous amino acid pools, as contrasted to a significant role
made by UP. Furthermore, there is also a question as to whether
L-glutamate dehydrogenase, located in the mitochondrial
marix, makes any real or significant net contribution to
NH4 assimilation in the animal system, because
the Km for NH4+ is
quite high (5–40 mmol/L; Katagari and Nakamura 1999, 1 mmol/L;
Lehninger et al. 1993). Another pathway of NH4+
assimilation could be via glycine synthase, and the glycine formed then
could be via serine hydroxymethyl transferase. However, serine
liberates its -amino N as NH4+, via the action of
serine dehydratase, and so the combined action of these three enzymes
would not result in a net accumulation of -amino N in the mixed
amino acid pools of the body. For these biochemical reasons, it appears
plausible that under normal conditons UP would be the more important
site for the regulation of body N loss. In this context, and as clearly
outlined by Waterlow (1999)
, there is still a great deal of uncertainty
as to the in vivo mechanisms responsible for both the short- and
longer-term regulation of UP under differing pathophysiological
circumstances. Urea synthesis is affected by pH, hormones and
substrates (Meijer et al. 1990
) and Nissim et al. (1996)
have suggested this might involve mechanisms including i)
alteration of citrulline synthesis, ii) formation of
aspartate from pyruvate via the pyruvate carboxylase pathway and/or
iii) modulation of N-acetyglutamate synthesis from
pyruvate, via the pyruvate dehydrogenase pathway. Finally, it must be
appreciated that there is a structural and functional organization of
hepatic ammonia metabolism, with different roles played by the
periportal (low-affinity system for urea synthesis) and perivenous
(high affinity system for glutamine synthesis) zones
(Haussinger1990
, Haussinger et al. 1992
).
This also makes it difficult to fully comprehend or predict the precise
fate of ammonia entering the portal circulation. A collaboration among
cellular/molecular studies and in vivo metabolic investigations of
metabolic N transfer would assist in reducing this gap in our knowledge
of the mechanisms responsible for body N homeostasis.
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FOOTNOTES |
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Manuscript received May 21, 1999. Initial review completed July 26, 1999. Revision accepted November 29, 1999.
|
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|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
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1. Child S. C., Soares M. J., Reed M., Persaud C., Forrester T., Jackson A. A. Urea kinetics varies in Jamaican women and men in relation to adiposity, lean body mass and protein intake. Europ. J. Clin. Nutr. 1997;51:107-115[Medline]
2. Danielsen M., Jackson A. A. Limits of adaptation to a diet low in protein in normal man: urea kinetics. Clin .Sci. 1992;83:103-108[Medline]
3.
El-Khoury A. E., Ajami A. M., Fukagawa N. K., Chapman T. E., Young V. R. Diurnal pattern of the interrelationships between leucine oxidation, urea production and hydrolysis in humans. Am. J. Physiol. 1996;271:E563-E573
4.
El-Khoury A. E., Basile A., Beaumie L., Wang S. Y., Al-Amiri H. A., Selvaraj A., Wong S., Atkinson A., Ajami A. M., Young V. R. Twenty-four hour intravenous and oral tracer studies with L-[1-13C]-2--amino adipic acid and L-[1-13C]lysine as tracers at generous nitrogen and lysine intakes in healthy adults. Am. J. Clin. Nutr. 1998;86:827-839
5.
El-Khoury A. E., Forslund A., Olsson R., Branth S., Sjodin A., Andersson A., Atkinson A., Selvaraj A., Hambraeus L., Young V. R. Moderate exercise at energy balance does not affect 24h leucine oxidation or nitrogen retention in healthy men. Am. J. Physiol. 1997;273:E394-E407
6.
El-Khoury A. E., Fukagawa N. K., Sánchez M., Tsay R. H., Gleason R. E., Chapman T. E., Young V. R. Validation of the tracer balance concept with reference to leucine: 24-h intravenous tracer studies with L-[1-13C]leucine and [15 N-15 N]urea. Am. J. Clin. Nutr. 1994;59:1000-1011
7. FNB Recommended Dietary Allowances (Subcommittee of the Tenth Edition of the RDAs, Food and Nutrition Board, Commission on Life Sciences, National Research Council) 10th edn. 1989:284 National Academy Press Washington, D.C.
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Forslund A. E., Hambraeus L., Olsson R. M., El-Khoury A. E., Yu Y.-M, Young V. R. The 24-h whole body leucine and urea kinetics at normal and high protein intakes with exercise in healthy adults. Am. J. Physiol. 1998;275:E310-E320
9. Giordano C., dePascale C., Balestrieri C., Cittodini D., Grescenzi A. Incorporation of urea 15 N in amino acids of patients with chronic renal failure on low nitrogen diet. Am. J. Clin. Nutr. 1968;21:394-404[Abstract]
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Hamadeh M. J., Hoffer L. J. Tracer methods underestimate short-term variations in urea production in humans. Am. J. Physiol. 1998;274:E547-E553
11. Haussinger D. Nitrogen metabolism in liver: Structural and functional organization and physiological relevance. Biochem. J. 1990;267:271-290
12. Haussinger D., Lamers W. H., Moorman A. F. M. Hepatocyte heterogeneity in the metabolism of amino acids and ammonia. Enzyme 1992;46:72-93[Medline]
13. Jackson A. A. Critique of protein-energy interactions in vivo: urea kinetics. Scrimshaw N. S. Schürch B. eds. Protein-Energy Interactions 1992:163-179 Nestlé Foundation Lausanne, Switzerland.
14. Jackson A.A. Salvage of urea-nitrogen in the large bowel: functional significance in metabolic control and adaptation. Biochem. Soc. Trans. 1998;26:231-236[Medline]
15. Jackson A. A. Limits of adaptation to high dietary protein intakes. Europ. J. Clin. Nutr. 1999;52(Suppl 1):S44-S52
16. Jackson A. A., Danielsen M. S., Boyes S. Noninvasive method for measuring urea kinetics with a single dose of [15 N15 N]urea in free-living humans. J. Nutr. 1993;123:2129-2136
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